title: The Yersinia pseudotuberculosis and Yersinia pestis toxin complex is active against cultured mammalian cells.
abstract: The toxin complex (Tc) genes were first identified in the insect pathogen Photorhabdus luminescens and encode approximately 1 MDa protein complexes which are toxic to insect pests. Subsequent genome sequencing projects have revealed the presence of tc orthologues in a range of bacterial pathogens known to be associated with insects. Interestingly, members of the mammalian-pathogenic yersiniae have also been shown to encode Tc orthologues. Studies in Yersinia enterocolitica have shown that divergent tc loci either encode insect-active toxins or play a role in colonization of the gut in gastroenteritis models of rats. So far little is known about the activity of the Tc proteins in the other mammalian-pathogenic yersiniae. Here we present work to suggest that Tc proteins in Yersinia pseudotuberculosis and Yersinia pestis are not insecticidal toxins but have evolved for mammalian pathogenicity. We show that Tc is secreted by Y. pseudotuberculosis strain IP32953 during growth in media at 28 degrees C and 37 degrees C. We also demonstrate that oral toxicity of strain IP32953 to Manduca sexta larvae is not due to Tc expression and that lysates of Escherichia coli BL21 expressing the Yersinia Tc proteins are not toxic to Sf9 insect cells but are toxic to cultured mammalian cell lines. Cell lysates of E. coli BL21 expressing the Y. pseudotuberculosis Tc proteins caused actin ruffles, vacuoles and multi-nucleation in cultured human gut cells (Caco-2); similar morphology was observed after application of a lysate of E. coli BL21 expressing the Y. pestis Tc proteins to mouse fibroblast NIH3T3 cells, but not Caco-2 cells. Finally, transient expression of the individual Tc proteins in Caco-2 and NIH3T3 cell lines reproduced the actin and nuclear rearrangement observed with the topical applications. Together these results add weight to the growing hypothesis that the Tc proteins in Y. pseudotuberculosis and Y. pestis have been adapted for mammalian pathogenicity. We further conclude that Tc proteins from Y. pseudotuberculosis and Y. pestis display differential mammalian cell specificity in their toxicity.
authors.affiliation: University of Exeter in Cornwall, Department of Biosciences, Tremough Campus, Penryn, Cornwall TR10 9EZ, UK.|University of Bath, Department of Biology and Biochemistry, Claverton Down, Bath BA2 7AY, UK.
authors.forename: Michelle C
authors.initials: MC
authors.lastname: Hares
authors.affiliation: London School of Hygiene and Tropical Medicine, London WC1E 7HT, UK.|University of Exeter in Cornwall, Department of Biosciences, Tremough Campus, Penryn, Cornwall TR10 9EZ, UK.
authors.forename: Stewart J
authors.initials: SJ
authors.lastname: Hinchliffe
authors.affiliation: London School of Hygiene and Tropical Medicine, London WC1E 7HT, UK.
authors.forename: Philippa C R
authors.initials: PCR
authors.lastname: Strong
authors.affiliation: CNRS-UPR9022, Institut de Biologie Moléculaire et Cellulaire, 15 rue René Descartes, 67084 Strasbourg Cedex, France.
authors.forename: Ioannis
authors.initials: I
authors.lastname: Eleftherianos
authors.affiliation: University of Exeter in Cornwall, Department of Biosciences, Tremough Campus, Penryn, Cornwall TR10 9EZ, UK.
authors.forename: Andrea J
authors.initials: AJ
authors.lastname: Dowling
authors.affiliation: University of Exeter in Cornwall, Department of Biosciences, Tremough Campus, Penryn, Cornwall TR10 9EZ, UK.
authors.forename: Richard H
authors.initials: RH
authors.lastname: Ffrench-Constant
authors.affiliation: University of Bath, Department of Biology and Biochemistry, Claverton Down, Bath BA2 7AY, UK.
authors.forename: Nick
authors.initials: N
authors.lastname: Waterfield
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D001427:Bacterial Toxins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D018938:Caco-2 Cells; D002460:Cell Line; D005347:Fibroblasts / Q000187:drug effects / Q000382:microbiology; D006801:Humans; D018402:Manduca / Q000382:microbiology; D051379:Mice; D041681:NIH 3T3 Cells; D021381:Protein Transport; D015009:Yersinia Infections / Q000382:microbiology*; D015010:Yersinia pestis / Q000235:genetics / Q000378:metabolism*; D015011:Yersinia pseudotuberculosis / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity*
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins